The long-term objectives of this application are to understand the molecular mechanisms of resistance to androgen-directed therapies and to develop effective strategies to overcome the resistance. Recent significant advances in our understanding of continued androgen receptor (AR) signaling in castration-resistant prostate cancer have led to the development and FDA approval of two next-generation androgen-directed therapies, abiraterone and enzalutamide (MDV3100), which heralded a new era of prostate cancer therapy. However, disease progression after androgen-directed therapies remains the most critical challenge in the clinical management of prostate cancer. Upregulated expression of constitutively-active, alternatively-spliced AR variants (AR-Vs) that lack the ligand binding-domain has been implicated to play an important role in disease progression. However, the precise mechanism by which androgen-directed therapies increase AR-V expression and the precise mechanism by which AR-Vs regulate target-gene expression in mediating therapeutic resistance remain poorly understood. These are two issues vital to understanding resistance to androgen-directed therapies and developing effective means to overcome the resistance, and therefore are the main focus of investigation of the proposed study. Based on the findings from ours and others, we propose that upregulation of AR-Vs is an inevitable response of prostate cancers to all androgen-directed therapies currently accepted in the clinic and that a shift from androgen-dependent full-length AR (AR-FL) homodimerization to androgen-independent AR-FL/AR-V and AR-V/AR-V dimerization leads to unleashed AR activity, driving disease progression. Given the potency of abiraterone and enzalutamide and the evidence for ongoing suppression of androgen synthesis at the time of progression on abiraterone, this resistance mechanism may be especially critical for abiraterone and enzalutamide. Three specific aims are proposed to test the hypotheses. Aim 1: Delineate the mechanism of AR-V induction following androgen-directed therapies. Aim 2: Elucidate the mechanism by which AR-Vs regulate target-gene expression. Aim 3: Correlate AR-V expression in circulating tumor cells with abiraterone/enzalutamide response. The proposed research is relevant to public health because it addresses a resistance mechanism that is inherent to all current androgen-directed therapies, including the new drugs abiraterone and enzalutamide. Moreover, we hope to establish AR-Vs as a prognostic marker to guide the selection of patients who will likely benefit from abiraterone and enzalutamide therapies. Overall, the outcome of this study is expected to have a direct impact on rational drug design and combination to overcome the shortcoming of current therapies and on reducing suffering and improving quality of life of the patients.